title: Potentialities of aberrantly methylated circulating DNA for diagnostics and post-treatment follow-up of lung cancer patients.
abstract: To date, aberrant DNA methylation has been shown to be one of the most common and early causes of malignant cell transformation and tumors of different localizations, including lung cancer. Cancer cell-specific methylated DNA has been found in the blood of cancer patients, indicating that cell-free DNA circulating in the blood (cirDNA) is a convenient tumor-associated DNA marker that can be used as a minimally invasive diagnostic test. In the current study, we investigated the methylation status in blood samples of 32 healthy donors and 60 lung cancer patients before and after treatment with neoadjuvant chemotherapy followed by total tumor resection. Using quantitative methylation-specific PCR, we found that the index of methylation (IM), calculated as IM = 100 × [copy number of methylated/(copy number of methylated + unmethylated gene)], for the RASSF1A and RARB2 genes in the cirDNA isolated from blood plasma and cell-surface-bound cirDNA was elevated 2- to 3-fold in lung cancer patients compared with healthy donors. Random forest classification tree model based on these variables combined (RARB2 and RASSF1A IM in both plasma and cell-surface-bound cirDNA) lead to NSCLC patients' and healthy subjects' differentiation with 87% sensitivity and 75% specificity. An association of increased IM values with an advanced stage of non-small-cell lung cancer was found for RARB2 but not for RASSF1A. Chemotherapy and total tumor resection resulted in a significant decrease in the IM for RARB2 and RASSF1A, in both cirDNA fractions, comparable to the IM level of healthy subjects. Importantly, a rise in the IM for RARB2 was detected in patients within the follow-up period, which manifested in disease relapse at 9 months, confirmed with instrumental and pathologic methods. Our data indicate that quantitative analysis of the methylation status of the RARB2 and RASSF1A tumor suppressor genes in both cirDNA fractions is a useful tool for lung cancer diagnostics, evaluation of cancer treatment efficiency and post-treatment monitoring.
authors.affiliation: Cancer Research Institute of Siberian Branch of the Russian Academy of Medical Sciences, Tomsk, Russia. Electronic address: anastasia-ponomaryova@rambler.ru.
authors.forename: Anastasia A
authors.initials: AA
authors.lastname: Ponomaryova
authors.affiliation: Institute of Chemical Biology and Fundamental Medicine of Siberian Branch of the Russian Academy of Sciences, Novosibirsk, Russia.
authors.forename: Elena Yu
authors.initials: EY
authors.lastname: Rykova
authors.affiliation: Cancer Research Institute of Siberian Branch of the Russian Academy of Medical Sciences, Tomsk, Russia; Siberian State Medical University, Tomsk, Russia.
authors.forename: Nadezda V
authors.initials: NV
authors.lastname: Cherdyntseva
authors.affiliation: Institute of Chemical Biology and Fundamental Medicine of Siberian Branch of the Russian Academy of Sciences, Novosibirsk, Russia.
authors.forename: Tatiana E
authors.initials: TE
authors.lastname: Skvortsova
authors.affiliation: Cancer Research Institute of Siberian Branch of the Russian Academy of Medical Sciences, Tomsk, Russia.
authors.forename: Alexey Yu
authors.initials: AY
authors.lastname: Dobrodeev
authors.affiliation: Cancer Research Institute of Siberian Branch of the Russian Academy of Medical Sciences, Tomsk, Russia.
authors.forename: Alexander A
authors.initials: AA
authors.lastname: Zav'yalov
authors.affiliation: Institute of Cytology and Genetics of Siberian Branch of the Russian Academy of Sciences, Novosibirsk, Russia.
authors.forename: Leonid O
authors.initials: LO
authors.lastname: Bryzgalov
authors.affiliation: Cancer Research Institute of Siberian Branch of the Russian Academy of Medical Sciences, Tomsk, Russia; Siberian State Medical University, Tomsk, Russia.
authors.forename: Sergey A
authors.initials: SA
authors.lastname: Tuzikov
authors.affiliation: Institute of Chemical Biology and Fundamental Medicine of Siberian Branch of the Russian Academy of Sciences, Novosibirsk, Russia.
authors.forename: Valentin V
authors.initials: VV
authors.lastname: Vlassov
authors.affiliation: Institute of Chemical Biology and Fundamental Medicine of Siberian Branch of the Russian Academy of Sciences, Novosibirsk, Russia.
authors.forename: Pavel P
authors.initials: PP
authors.lastname: Laktionov
mesh_terms: D000328:Adult; D000368:Aged; D002289:Carcinoma, Non-Small-Cell Lung / Q000175:diagnosis* / Q000235:genetics* / Q000628:therapy; D004247:DNA / Q000097:blood*; D019175:DNA Methylation; D005260:Female; D005500:Follow-Up Studies; D006801:Humans; D008175:Lung Neoplasms / Q000175:diagnosis* / Q000235:genetics* / Q000628:therapy; D008297:Male; D008875:Middle Aged; D009367:Neoplasm Staging; D025341:Principal Component Analysis; D011379:Prognosis; D018168:Receptors, Retinoic Acid / Q000235:genetics; D012307:Risk Factors; D012680:Sensitivity and Specificity; D025521:Tumor Suppressor Proteins / Q000235:genetics
chemical_list: C533416:RARB2 protein, human; C411762:RASSF1 protein, human; D018168:Receptors, Retinoic Acid; D025521:Tumor Suppressor Proteins; D004247:DNA
keywords: Circulating DNA; Diagnostics; Lung cancer; Methylation; Monitoring; Prognosis; Tumor suppressor gene